Project Abstract This project proposes to study how connections between nerve cells in the C. elegans brain process information to generate behavior. Healthy brain function is critically important to our everyday lives, and it is the connections among nerve cells (neurons) that confer the brain its ability to perceive the environment and to control our actions. Similarly, changes to the patterns and properties of neural connections can dramatically alter brain function and have been implicated in aging-related cognitive decline and in disease. For this reason, neuroscientists have long sought to characterize neural connections in the brain. And because many cognitive disorders and diseases are not restricted to one region of the brain, but are instead global brain phenomenon, tools are needed to characterize neural connections across the entire brain. But just knowing who is connected to whom is not enough. Rather, it is the strength and sign of the connections between neurons as well as their excitability and temporal properties that ultimately determine how a collection of neurons process information and generate behavior. Functional connectivity is the term used to describe the detailed properties of neural connections in the brain, and in contrast to a wiring diagram or `connectome,' a map of functional connectivity captures the details of how each neuron's activity affects others in the network. Currently no method exists that provides the combined resolution and scale necessary to directly measure functional connectivity at cellular resolution of an entire brain for any animal, and especially not during unrestrained behavior. A major hurdle has been the lack of a tractable model system in which to develop brain-wide probes of functional connectivity. To overcome this hurdle, I propose to work in the small nematode C. elegans. In contrast to mammalian brains, C. elegans has a compact nervous system of only 302 neurons and is the only organism to have a complete map of its neuroanatomical wiring. I will develop new techniques for measuring and interpreting whole-brain functional connectivity in the nematode C. elegans at cellular resolution during unrestrained behavior. By innovating new tools for the worm, I will develop solutions to technical challenges now, that will later be translated to vertebrate systems. To measure brain-wide functional connectivity, I will develop a new optical neurophysiology microscope to sequentially activate each neuron in the brain, while simultaneously recording activity from every neuron. I will infer synaptic strength and neural excitability from the network's response. I will perform these measurements in freely moving animals to study the interplay between functional connectivity and animal behavior. And I will apply measurements of functional connectivity to address outstanding questions about how neural circuits in C. elegans change due to the animal's behavior state, learning, or aging. I will also directly compare the animal's anatomical wiring, or connectome, to its functional connectivity. By measuring the functional connections in the brain of a moving animal I will investigate fundamental questions that relate brain structure, function and organism behavior.